Phosphatidylcholine is an important constituent of eukaryotic cell membranes, secreted lipoproteins, and signal transduction pathways. Its biosynthesis is regulated in part by the first enzyme of the biosynthetic pathway for phosphatidylcholine, choline kinase. The objective of the research outlined in this proposal is to understand the mechanism of regulation of choline kinase activity. The hypothesis to be tested is that the regulation involves differential expression of multiple isozymes of choline kinase, some of which are generated by alternative splicing and others are encoded in separate genes. These studies will focus on two organisms. To extend previous studies on mammalian choline kinase, mouse tissues and cell lines will be used. To begin to use a molecular genetic approach for studying these enzymes, the invertebrate C. elegans will be used. The specific aims are I) To develop molecular tools to investigate murine choline kinase. These tools will include cDNAs, antibodies, and recombinant protein. 2) To investigate patterns of expression of choline kinase isozymes in adult and embryonic mouse tissues and mouse cell lines expressing the ras oncogene. 3) To identify and characterize murine choline kinase genes. 4) To express and characterize putative choline kinase isozymes of C. elegans. 5) To investigate patterns of choline kinase expression in C. elegans. The proposed research will provide fundamental knowledge about how the synthesis of phosphatidylcholine is regulated, and about how choline kinase may be involved in other cellular functions.